Legal Medicine - Articles in Press

Quantitative analysis of GFAP- and S100 protein-immunopositive astrocytes to investigate the severity of traumatic brain injury - Corrected Proof

2012-02-02

Abstract: Previous studies have shown that diffuse cortical astrocyte damage is seen in acute deaths due to brain injury and mechanical asphyxiation. The present study quantitatively investigated the number of astrocytes that showed GFAP- and S100-protein immunopositivity in the cerebral white matter and hippocampus at the sites distant from primary injury with regard to survival time, complication, and the immediate cause of death of brain injury cases. Autopsy cases of brain injury (8–48h postmortem) comprising acute/subacute deaths (survival time, <3/6h–3days; n=27/42) and delayed deaths (survival time >3days) with/without complications (n=30/22) were examined. Delayed death cases with complications were subdivided into those in which the immediate cause of death had been determined as cerebral dysfunction (n=22) and those that had been determined as due to fatal complications (n=8). For controls, natural deaths from pneumonias (n=12) and sudden cardiac deaths (n=27) were used. In brain injury cases, the numbers of astrocytes in the cerebral white matter and hippocampal CA4 region were significantly lower for subacute death and delayed death without complications (p<0.05–0.001). Delayed death with fatal complications showed a significant increase in the number of astrocytes (p<0.05). Among delayed death cases, the numbers of astrocytes were higher in the cases with fatal complications than in those without complications and with non-fatal complications, although the latter cases showed large variations in the numbers of these astrocytes. These findings suggest that critical brain injury causes acute death without evident astrocyte pathology and that subacute death is associated with progressive brain damage accompanied by an astrocyte loss. In delayed death cases, the numbers astrocytes might be closely related to the severity of posttraumatic brain injury. GFAP and S100-immunopositivity might be useful for elucidating the cause and process of deaths due to brain injury.

Forensic histopathology: Fundamentals and perspectives - Corrected Proof

Toshikazu Kondo — 2012-02-01

On 1984, Prof. Werner Jannsen published a monograph “Forensic Histopathology”, which was the first comprehensive textbook of forensic pathology. At that time, lots of forensic pathologists were deeply impressed that the book included wealth information with instructive illustrations. It is necessary for forensic pathologists to catch up with the novel knowledges in order to correctly diagnose the cause of death and to explain the mechanisms of violence. Thus, many forensic pathologists are looking forward to a new textbook of forensic histopathology. On 2011 a new book, “Forensic Histopathology: Fundamentals and Perspectives”, providing up-to-date knowledges on forensic histopathology to forensic pathologists as well as general pathologists, has been published by Prof. Reinhard B. Dettmeyer. He is most suitable forensic pathologist for publishing forensic histopathology, because he is a specialist in both pathology and forensic pathology. It is very honor for me to give me an opportunity for writing this book review.

The effects of low-dose methamphetamine pretreatment on endoplasmic reticulum stress and methamphetamine neurotoxicity in the rat midbrain - Corrected Proof

Toshiaki Takeichi, Elaine Lu Wang, Osamu Kitamura — 2012-02-01

Abstract: Methamphetamine (METH) neurotoxicity is involved in METH-related deaths. It has been suggested that the midbrain, together with the striatum, is affected by METH neurotoxicity and the endoplasmic reticulum (ER) stress is induced in the processes of METH neurotoxicity. In this study, we examined the effects of low-dose METH administration for 5d on GRP78 and C/EBP homologous protein (CHOP), both of which are induced under ER stress, and METH neurotoxicity in the rat midbrain. We showed that 1mg/kg of METH induced an increase in GRP78 protein and mRNA expression 1d after the last injection, but had no effect on the levels of CHOP, tyrosine hydroxylase (TH), or GFAP. Secondly, we evaluated the induction of ER stress and the extent of METH neurotoxicity in the midbrain of animals pretreated with METH. In animals pretreated with saline, we observed elevated CHOP levels, together with decreased TH levels and increased GFAP levels, indicative of METH neurotoxicity, after neurotoxic METH administration, while there was no significant change in GRP78 levels. In contrast, low-dose METH (1.0mg/kg) pretreatment increased GRP78 levels and inhibited the induction of CHOP in the midbrain without METH neurotoxicity. These findings of ER stress in animals pretreated with METH were associated with an early increase in SOD1 levels and upregulation of Bcl-2. Therefore, our study suggests that pretreatment with low-dose METH may be protective against METH neurotoxicity in the midbrain, leading to the suppression of oxidative stress and apoptotic mechanisms, in part via ER stress-related pathways. Because chronic human METH abusers administrate low-dose METH repeatedly over an extended period before lethal injection, investigation of the pathophysiology of METH neurotoxicity in animals pretreated with low-dose METH might provide useful information on the pathophysiology of chronic and/or lethal METH use in cases of METH-related deaths.

The post mortem temperature plateau and its role in the estimation of time of death. A review - Corrected Proof

Jimmy L. Smart, Michał Kaliszan — 2012-01-30

Abstract: The purpose of this paper was to examine evidence to seek an explanation of the possible cause(s) or contributing factors to the temperature plateau phenomenon and its influence on time of death (TOD) estimation. The concept of the temperature plateau effect (TPE) is reviewed, and investigation is conducted into its possible prediction under post mortem conditions. The conclusion of this paper is that the appearance of a TPE in postmortem body core temperature decay curves is currently random and cannot be predicted. This unpredictability is based upon the interindividual differences in states (core body temperature, hyperthermia, use of drugs, trauma, etc.) and biomarker concentrations (electrolytes, thyroxine, etc.) at antemortem times, which will ultimately affect the shape of the postmortem temperature decay curve. However, studies indicated that the TPE is diminished or even absent in the head tissues, including eye and ear. The possibility of precise estimation of the TOD in the early post mortem period based on eye temperature measurements is also commented.

Molecular biological analysis of cardiac effect of high temperature in rats - Corrected Proof

Yasuhisa Nakagawa, Hiromasa Inoue, Kotaro Shinone, Mayumi Ikemura, Masayuki Nata — 2012-01-30

Abstract: The aim of this study was to investigate direct effects of heat exposure on the heart molecular-biologically and pathohistologically, using rats exposed to high temperatures. The mRNA expression of natriuretic peptide type A (Nppa), natriuretic peptide type B (Nppb), actin alpha 1 skeletal muscle (Acta1), myosin heavy polypeptide 6 cardiac muscle alpha (Myh6) and myosin heavy polypeptide 7 cardiac muscle alpha (Myh7) was determined in the hearts of the rats. Whereas the expression of Nppa and Nppb rapidly increased immediately after the heat exposure, the expression of Acta1 was gradually reduced, which indicated cardiac overload. Moreover, the expression of Myh6 and Myh7 in the heart increased 4h after the heat exposure, which suggested the involvement of a compensatory mechanism. Immunohistochemical staining with anti-fibronectin antibody showed that positive cardiomyocytes could be detected sparsely 4h after the heat exposure, and they could be clearly observed 8h after the heat exposure. Our results showed that hyperthermia causes myocardial damage shortly after the exposure to heat and that the ventricle was more vulnerable to hyperthermia-induced damage than the atrium. Cardiac dysfunction may be induced not only by hypercytokinemia but also by the direct effect of heat exposure at the early period of heat stroke, which may be one of the mechanisms by which heat causes death. Elucidating the mechanism of death from heat stroke could lead to not only diagnostic improvement but also the prevention of death from heat stroke.

Y chromosome STR allelic and haplotype diversity in a Rwanda population from East Central Africa - Corrected Proof

Kuppareddi Balamurugan, George Duncan — 2012-01-30

Abstract: We have analyzed 17 Y-chromosomal STR loci in a population sample of 69 unrelated male individuals of the Rwanda-Hutu population from East Central Africa using an AmpFlSTR® Yfiler™ PCR amplification kit. A total of 62 unique haplotypes were identified among the 69 individuals studied. The haplotype diversity was found to be 0.9970 for this population. The gene diversity ranged from 0.1130 (DYS392) to 0.7722 (DYS385). Comparison of populations in this study with twenty-five other national and global populations using Principal Co-ordinate Analysis (PCA) and phylogenetic molecular analysis using a genetic distance matrix indicates a delineation of all the African populations from other unrelated populations. The results of population pair-wise Fst p values indicate statistically significant differentiation of the Rwandan population when compared with 25 other global populations including four African populations (p=0.0000). Analysis of Molecular Variance (AMOVA) of the Rwanda population with four other African populations indicated a 93% variance within populations and 7% variance among the five populations. A data base search of the 62 haplotypes yielded only one non-African haplotype match, suggesting these haplotypes are unique to the African continent.

Verification of eye and skin color predictors in various populations - Corrected Proof

2012-01-30

Abstract: Validation of testing methods is an essential feature in all scientific endeavors, but it is particularly important in forensics. Due to the sensitive nature of these investigations and the limited sample size it is crucial to validate all employed procedures. This includes novel forensic phenotypic DNA tests, to learn more of their capabilities and limitations before incorporating them as routine methods. Ideally, validations are performed on large sample sets that mimic real cases.Recently, three phenotypic predictors, two for eye colors and one for skin color have been published (Spichenok et al., 2011; Walsh et al., 2011). These predictors are well-defined by a selection of single nucleotide polymorphisms (SNPs) and unambiguous instructions on how to interpret the genotypes. These standardized approaches have the advantages that they can be applied in diverse laboratories leading to the same outcome and offer the opportunity for validation. For these tests to be used on the characterization of human remains, they should be validated on various populations to perform reliably without prior knowledge of ethnic origin.Here, in this study, these eye and skin color predictors were validated on new sample sets and it could be confirmed that they can be applied in various populations, including African-American, South Asian (dark), East Asian (light), European, and mixed populations. The outputs were either predictive or inconclusive. Predictions were then compared against the actual eye and skin colors of the tested individuals. The error-rates varied; they were low for the predictors that describe the eye and skin color exclusively (non-brown or non-blue and non-white or non-dark, respectively) and higher for the predictor that describes individual eye colors (blue, brown, and intermediate/green), because of uncertainties with the green eye color prediction. Our investigation deepens the insight for these predictors and adds new information.

Diagnosis of aortic dextroposition on human skeletal remains - Corrected Proof

2012-01-30

Abstract: The fine macroscopic observation of a young adult female skeleton recovered from a Roman graveyard in Romania revealed distinctive flattening of the vertebra related to a right-sided aorta. Associated bone anomalies may be related to a Kartagener syndrome.This case highlights the fact that visceral anomalies may be diagnosed even on skeletal remains. Such lesions could be useful for osteo-archaeologists, of course, but also for forensic anthropologist investigators dealing with skeletonized remains (for example during the identification process of a dead body, through comparison with known medical data for missing people). More, hypotheses about cause and/or manner of death may be given, and a possibility of genetic confirmation exists.

Human blood identification using the genome profiling method - Corrected Proof

2012-01-30

Abstract: In criminal investigations, usually it is necessary to identify whether blood spots found at crime scenes are from humans or not. Nowadays, immunohistochemical methods and DNA analysis are usually used for this purpose. However, such methods and DNA analysis are labor intensive and expensive, and require highly trained skilled technicians. Recently, the genome profiling method (GP method) was developed. However, its use as a human DNA analysis method has not been reported. In this report, an attempt was made to differentiate human blood samples from animal blood samples using the GP method for forensic purposes.DNA extracted from a rat, squirrel, cat, dog, cow, and antelope along with human blood samples were analyzed. Following cluster analysis the human samples clustered into a single group separate from the animal samples. Therefore, although the number of samples was small the results suggest that the GP method might enable us to differentiate human samples from various animal samples. It may become a powerful tool in the field of forensic science.

Rapid and reliable screening method for detection of 70 pesticides in whole blood by gas chromatography–mass spectrometry using a constructed calibration-locking database - Corrected Proof

2012-01-30

Abstract: Pesticide poisoning is one of the most common causes of death by poisoning in Japan, and various kinds of pesticides including organophosphates, carbamates and pyrethroids are listed as causative substances. The purpose of our study was to develop a rapid and reliable screening method for various kinds of pesticides in whole blood by using a unique calibration-locking database and gas chromatography–mass spectrometry. A database of 70 pesticides was constructed using NAGINATA™ software with parameters such as mass spectrum, retention time and qualifier ion/target ion ratio (QT ratio) and calibration curve. Diazepam-d5 was used as the internal standard for construction of each calibration curve within the range of 0.01–5.0μg/ml. We examined the applicability of the constructed database by analyzing whole blood samples spiked with 70 pesticides. The pesticides in blood were extracted with hexane under acidic conditions or with an enhanced polymer column (Focus™), subjected to GC–MS, and screened by the pesticides database. Among the 70 pesticides examined, 66 and 62 were successfully identified at the level of 1 and 0.1μg/ml, respectively, by hexane and 63 and 51 were identified by the Focus column without the use of standard compounds. The time required for data analysis was significantly reduced. Since the established method can produce qualitative and semi-quantitative data without the need for standard substances, this new screening method using NAGINATA™ should be useful for confirming the presence of pesticides in blood in future clinical and forensic cases.